Multiplexed bioluminescence microscopy via phasor analysis

Imaging tools allow us to visualize biological processes, including single cell dynamics and metabolic fluxes. Tracking multiple cells or features over extended periods of time, though, remains challenging. Common approaches rely on fluorescent probes, which require intense excitation light that can damage cells and cause phototoxicity. Bioluminescence imaging (BLI) relies on light production from a chemical reaction, the luciferase-catalyzed oxidation of small molecule substrates (luciferins), and can offer a more bio-friendly alternative for monitoring cellular events in real time.